The objective of this proposal is to isolate a mammalian gene of known function, to characterize its physical organization, and to study its replication at a molecular level. The Chinese hamster ribosomal genes were selected because 1) they occur in multiple copies; 2) the initial enrichment step in the isolation of Chinese hamster ribosomal DNA has been worked out in preliminary experiments; 3) the time at which the ribosomal genes of synchronized Chinese hamster V79 tissue culture cells replicate during the cell cycle has been localized to the early portion of the S phase. Isolation and purification of these genes will be accomplished by equilibrium density centrifugation in Hg ions/Cs2SO4, actinomycin D-CsCl and neutral CsCl gradients. The ribosomal DNA will be characterized by thermal denaturation and renaturation studies, buoyant density, cleavage by restriction endonucleases and electron microscopy of partially denatured ribosomal DNA molecules. Replicating ribosomal DNA molecules will be prepared from DNA of synchronized cells and will be studied by electron microscopy of native and partially denatured ribosomal DNA.